Interferon-induced alteration of cellular factors affecting HIV alternative splicing, nuclear mRNA export and LTR transcription

人类免疫缺陷病毒1型（HIV-1）具有相对较小的基因组，其高度分散的顺式调控元件允许在选择性前mrna剪接水平上对其基因产物进行精确调控。由于选择性剪接是由细胞剪接机制处理的，细胞剪接调节蛋白，即异质核核糖核蛋白（hnRNP）和富含丝氨酸精氨酸的剪接因子（SRSF），对病毒复制至关重要。此外，某些hnRNP蛋白被证明参与细胞HIV mrna的核输出。此外，SRSF1或选定的hnRNPs的过表达与HIV ltr启动子的转录抑制有关。I型干扰素（ifn）通过诱导ifn刺激基因（ISG）的转录（如限制性因子APOBEC3G）在宿主细胞中建立抗病毒状态，在对抗HIV-1的先天免疫防御中发挥重要作用。我们可以证明，在临床使用的IFNα亚型2治疗后，T细胞和巨噬细胞中不同hnRNP和SRSF编码mrna的表达水平被特异性抑制，而IFNα14治疗的程度更高，IFNα14是抗HIV-1最有效的亚型。根据这些未发表的发现，对于hnRNP家族和SRSF蛋白，ifn抑制基因IRepGs似乎更合适，并描述了一种新的ifn诱导的影响hiv复制的分子机制。此外，我们的先例数据表明，干扰素诱导的抑制在hiv感染后被废除。因此，我们假设hnRNP和SRSF与HIV基因组中的顺式调控元件相互作用最有可能显著促进ifn对HIV的抗病毒作用。因此，在本提案中，我们的目标是研究I-III型ifn对细胞和病毒剪接、LTR转录和mRNA输出水平的影响，以及其对细胞系和患者源性原代细胞中HIV-1复制的影响。

The human immunodeficiency virus type 1 (HIV-1) has a relatively small genome, which is highly dispersed with cis-regulatory elements allowing a precise regulation of its gene products on the level of alternative pre-mRNA splicing. Since alternative splicing is processed by the cellular splicing machinery, cellular splicing regulatory proteins, which are heterogeneous nuclear ribonucleoproteins (hnRNP) and serine-arginine-rich splicing factors (SRSF), are crucial for viral replication. In addition, certain hnRNP proteins were shown to be involved in nuclear export of cellular HIV mRNAs. Furthermore, overexpression of SRSF1 or selected hnRNPs have been related to transcriptional inhibition of the HIV LTR-promoter. Type I Interferons (IFNs) play an important role in the innate immune defence against HIV-1 by inducing the transcription of IFN-stimulated genes (ISG) such as the restriction factor APOBEC3G establishing an anti-viral state in host cells. We could show that the expression levels of distinct hnRNP and SRSF coding mRNAs in T cells and macrophages are specifically repressed upon treatment with the clinically used IFNα subtype 2 and to an even higher extent with IFNα14, which is the most potent subtype against HIV-1. According to these unpublished findings, for the hnRNP family and SRSF proteins the term of IFN-repressed genes IRepGs seems to be more appropriate and describes a novel IFN-induced molecular mechanism affecting HIV-replication. Additionally, our precedent data demonstrate that in the interferon induced repression is abolished upon HIV-infection.We therefore assume that hnRNP and SRSF interacting with cis-regulatory elements in the HIV genome most likely significantly contribute to the anti-viral effect of IFNs against HIV. Thus, in this proposal we aim to investigate the impact of type I-III IFNs on the level of cellular and viral splicing, LTR transcription, and mRNA export, as well as its consequences for HIV-1 replication in cell lines and patient derived primary cells.